Mineo Kondo

TRPM1 is a component of the retinal ON bipolar cell transduction channel in the mGluR6 cascade

An essential step in intricate visual processing is the segregation of visual signals into ON and OFF pathways by retinal bipolar cells (BCs). Glutamate released from photoreceptors modulates the photoresponse of ON BCs via metabotropic glutamate receptor 6 (mGluR6) and G protein (Go) that regulates a cation channel. However, the cation channel has not yet been unequivocally identified. Here, we report a mouse TRPM1 long form (TRPM1-L) as the cation channel. We found that TRPM1-L localization is developmentally restricted to the dendritic tips of ON BCs in colocalization with mGluR6. TRPM1 null mutant mice completely lose the photoresponse of ON BCs but not that of OFF BCs. In the TRPM1-L-expressing cells, TRPM1-L functions as a constitutively active nonselective cation channel and its activity is negatively regulated by Go in the mGluR6 cascade. These results demonstrate that TRPM1-L is a component of the ON BC transduction channel downstream of mGluR6 in ON BCs.

Pikachurin, a dystroglycan ligand, is essential for photoreceptor ribbon synapse formation

Exquisitely precise synapse formation is crucial for the mammalian CNS to function correctly. Retinal photoreceptors transfer information to bipolar and horizontal cells at a specialized synapse, the ribbon synapse. We identified pikachurin, an extracellular matrix–like retinal protein, and observed that it localized to the synaptic cleft in the photoreceptor ribbon synapse. Pikachurin null-mutant mice showed improper apposition of the bipolar cell dendritic tips to the photoreceptor ribbon synapses, resulting in alterations in synaptic signal transmission and visual function. Pikachurin colocalized with both dystrophin and dystroglycan at the ribbon synapses. Furthermore, we observed direct biochemical interactions between pikachurin and dystroglycan. Together, our results identify pikachurin as a dystroglycan-interacting protein and demonstrate that it has an essential role in the precise interactions between the photoreceptor ribbon synapse and the bipolar dendrites. This may also advance our understanding of the molecular mechanisms underlying the retinal electrophysiological abnormalities observed in muscular dystrophy patients.

ISCEV guidelines for clinical multifocal electroretinography (2007 edition).

The clinical multifocal electroretinogram (mfERG) is an electrophysiological test of local retinal function. With this technique, many local ERG responses, typically 61 or 103, are recorded from the cone-driven retina under light-adapted conditions. This document specifies guidelines for performance of the test. It also provides detailed guidance on technical and practical issues, as well as on reporting test results. The main objective of the guidelines is to promote consistent quality of mfERG testing and reporting within and among centers. These 2007 guidelines, from the International Society for Clinical Electrophysiology of Vision (ISCEV: http://www.iscev.org), replace the ISCEV guidelines for the mfERG published in 2003.

Functional Roles of Otx2 Transcription Factor in Postnatal Mouse Retinal Development

ABSTRACT We previously reported that Otx2 is essential for photoreceptor cell fate determination; however, the functional role of Otx2 in postnatal retinal development is still unclear although it has been reported to be expressed in retinal bipolar cells and photoreceptors at postnatal stages. In this study, we first examined the roles of Otx2 in the terminal differentiation of photoreceptors by analyzing Otx2; Crx double-knockout mice. In Otx2+/−; Crx−/− retinas, photoreceptor degeneration and downregulation of photoreceptor-specific genes were much more prominent than in Crx−/− retinas, suggesting that Otx2 has a role in the terminal differentiation of the photoreceptors. Moreover, bipolar cells decreased in the Otx2+/−; Crx−/− retina, suggesting that Otx2 is also involved in retinal bipolar-cell development. To further investigate the role of Otx2 in bipolar-cell development, we generated a postnatal bipolar-cell-specific Otx2 conditional-knockout mouse line. Immunohistochemical analysis of this line showed that the expression of protein kinase C, a marker of mature bipolar cells, was significantly downregulated in the retina. Electroretinograms revealed that the electrophysiological function of retinal bipolar cells was impaired as a result of Otx2 ablation. These data suggest that Otx2 plays a functional role in the maturation of retinal photoreceptor and bipolar cells.

Negative regulation of ciliary length by ciliary male germ cell-associated kinase (Mak) is required for retinal photoreceptor survival

Cilia function as cell sensors in many organs, and their disorders are referred to as “ciliopathies.” Although ciliary components and transport machinery have been well studied, regulatory mechanisms of ciliary formation and maintenance are poorly understood. Here we show that male germ cell-associated kinase (Mak) regulates retinal photoreceptor ciliary length and subcompartmentalization. Mak was localized both in the connecting cilia and outer-segment axonemes of photoreceptor cells. In the Mak-null retina, photoreceptors exhibit elongated cilia and progressive degeneration. We observed accumulation of intraflagellar transport 88 (IFT88) and IFT57, expansion of kinesin family member 3A (Kif3a), and acetylated α-tubulin signals in the Mak-null photoreceptor cilia. We found abnormal rhodopsin accumulation in the Mak-null photoreceptor cell bodies at postnatal day 14. In addition, overexpression of retinitis pigmentosa 1 (RP1), a microtubule-associated protein localized in outer-segment axonemes, induced ciliary elongation, and Mak coexpression rescued excessive ciliary elongation by RP1. The RP1 N-terminal portion induces ciliary elongation and increased intensity of acetylated α-tubulin labeling in the cells and is phosphorylated by Mak. These results suggest that Mak is essential for the regulation of ciliary length and is required for the long-term survival of photoreceptors.

Retinal Dysfunction in Basigin Deficiency

PURPOSE To examine the retina of basigin (Bsg) knockout mice by electrophysiological and histologic methods and thereby to determine the possible function of Bsg in phototransduction and retinal development. METHODS Scotopic and photopic electroretinograms (ERGs) were recorded from 11 wild-type, 12 heterozygous, and 8 homozygous Bsg gene knockout mice of different ages. The retinas were also examined by histologic and immunolabeling methods. RESULTS Bsg knockout mice of 5 to 41 weeks of age showed a decrease in the amplitude of all components of both the photopic and scotopic ERGs. In contrast, the fundus and the fluorescein fundus angiography and morphology of the retina at the light microscopic level appeared to be normal until 8 weeks of age in Bsg knockout mice. Thereafter, the length of outer segment and outer nuclear layers decreased with increasing age. Immunohistochemical analysis localized Bsg protein in a variety of cells in the retina, especially in the pigment epithelium, the upper outer plexiform layer and the inner segments of photoreceptor cells. CONCLUSIONS The results demonstrated that both rod and cone function were severely affected from an early age by the targeted disruption of the Bsg gene. In spite of abnormal ERGs, the photoreceptor cells maintained normal morphology up to 8 weeks. Thereafter, the photoreceptor cells degenerated gradually and were almost ablated by 41 weeks.

Intravitreal injection of bevacizumab for macular edema secondary to branch retinal vein occlusion:results after 12 months and multiple regression analysis.

Purpose: To evaluate the 12-month follow-up results of intravitreal bevacizumab therapy for macular edema secondary to branch retinal vein occlusion and to identify the pretreatment factors that were associated with an improvement of the final visual outcome. Methods: Fifty eyes of 50 patients with macular edema secondary to branch retinal vein occlusion received an injection of 1.25 mg/0.05 mL bevacizumab. Additional injections were done when recurrence of macular edema occurred or the treatment was not effective. The best-corrected visual acuity and foveal thickness were measured. Stepwise multiple regression analyses were also performed. Results: The mean logarithm of the minimum angle of resolution visual acuity improved significantly from 0.53 to 0.26, and the mean foveal thickness decreased significantly from 523 to 305 &mgr;m during the 12-month follow-up period. The mean number of injections was 2.0 (range, 1–4). Stepwise multiple regression analyses showed that younger patients had both better visual acuity at 12 months and greater improvement of visual acuity during 12 months. In addition, better pretreatment visual acuity was associated with better visual acuity at 12 months but with less improvement of the visual acuity. Conclusion: Intravitreal bevacizumab therapy can be a long-term effective treatment for macular edema secondary to branch retinal vein occlusion.

Blimp1 Suppresses Chx10 Expression in Differentiating Retinal Photoreceptor Precursors to Ensure Proper Photoreceptor Development

The zinc finger transcription factor Blimp1 plays fundamentally important roles in many cell lineages and in the early development of several cell types, including B and T lymphocytes and germ cells. Although Blimp1 expression in developing retinal photoreceptor cells has been reported, its function remains unclear. We identified Blimp1 as a downstream factor of Otx2, which plays an essential role in photoreceptor cell fate determination. To investigate Blimp1 function in the mouse retina, we ablated Blimp1 in the developing retina by conditional gene targeting. In the Blimp1 conditional knockout (CKO) retina, the number of photoreceptor cells was markedly reduced in the differentiated retina. We found that the numbers of both bipolar-like cells and proliferating retinal cells increased noticeably, with ectopic localizations in the postnatal developing retina. In contrast, a reduction of the number of photoreceptor precursors was observed during development. Forced expression of Blimp1 by in vivo electroporation suppressed bipolar cell genesis in the developing retina. Multiple genes involved in bipolar development, including Chx10, were upregulated in the Blimp1 CKO retina. Furthermore, we showed that Blimp1 can bind to the Chx10 enhancer and repress Chx10 enhancer activity. These results suggest that Blimp1 plays a crucial role in photoreceptor development by repressing genes involved in bipolar cell fate specification and retinal cell proliferation in differentiating photoreceptor precursors.

Multifocal electroretinogram in occult macular dystrophy

PURPOSE Occult macular dystrophy (OMD) is an unusual macular dystrophy presenting with an essentially normal fundus and fluorescein angiography but with progressive central visual loss. The authors studied the function of local retinal areas in the posterior pole of patients with OMD using multifocal electroretinograms (ERGs). METHODS Multifocal ERGs were recorded using the Visual Evoked Response Imaging System with 61 hexagonal elements within a visual field of 30 degrees radius from 8 OMD patients and 20 age-matched, normal subjects. The amplitudes and implicit times of the patients and normal control subjects were compared at the various retinal eccentricities. RESULTs. The amplitudes of the multifocal ERGs in the OMD patients were markedly reduced in the central 7 degrees of the fovea. The difference of the ERG amplitudes between OMD and normal subjects became smaller toward the peripheral retina. Most OMD patients had slight but significantly delayed implicit times across the whole testing field, and the differences between the OMD and the normal subjects did not change with retinal eccentricity. CONCLUSIONS Our results for multifocal ERG amplitudes support the idea that OMD patients have localized retinal dysfunction distal to the ganglion cells in the central retina. The delayed implicit times across the whole test field suggest that the retinal dysfunction has a broader boundary than expected by ERG amplitudes and psychophysical perimetric results.

Retinal remodeling in the Tg P347L rabbit, a large-eye model of retinal degeneration.

Retinitis pigmentosa (RP) is an inherited blinding disease characterized by progressive loss of retinal photoreceptors. There are numerous rodent models of retinal degeneration, but most are poor platforms for interventions that will translate into clinical practice. The rabbit possesses a number of desirable qualities for a model of retinal disease including a large eye and an existing and substantial knowledge base in retinal circuitry, anatomy, and ophthalmology. We have analyzed degeneration, remodeling, and reprogramming in a rabbit model of retinal degeneration, expressing a rhodopsin proline 347 to leucine transgene in a TgP347L rabbit as a powerful model to study the pathophysiology and treatment of retinal degeneration. We show that disease progression in the TgP347L rabbit closely tracks human cone‐sparing RP, including the cone‐associated preservation of bipolar cell signaling and triggering of reprogramming. The relatively fast disease progression makes the TgP347L rabbit an excellent model for gene therapy, cell biological intervention, progenitor cell transplantation, surgical interventions, and bionic prosthetic studies. J. Comp. Neurol. 519:2713–2733, 2011.